The short-term goal of this project is to utilize a proteomics approach to define the timing and stages of the onset of puberty. The study will utilize urine and blood samples collected previously from girls between the ages of 10-13 in an NCI-funded U54 observational study. Specimens were obtained from pre-menarcheal girls at study entry and at the following 12 and 24 months. Dietary questionnaires were taken at each time-point. Monthly questionnaires and telephone interviews were performed to assess the development of puberty. The long-term goal of this study is to better understand the biochemical and physiological events that occur during puberty since a study in genetically identical twins strongly suggested that the onset of puberty was controlled by external environmental effects, not genetic factors. In addition, the study found that earlier the onset of puberty, the higher the adult risk of breast cancer. The hypothesis is that puberty causes marked changes in the expression of steroid-sensitive genes and hence their products, the proteins. Since most cell lifetimes are relatively short (days-weeks), the proteins in them are subjected to turnover and will appear in blood as peptides. In addition, because of the large-scale redistribution of the body during puberty, peptides from proteins in non-estrogen sensitive tissues will also undergo significant changes. Although urine contains some intact proteins, filtration by kidney glomeruli results in the bulk of detectable peptides being below 20 kDa. The peptides will be recovered from urine using a high throughput hydrophobic cartridge and then eluted to yield a predominantly peptide fractionIn this study, we will (1) first use MALDI-TOF MS to establish potential peptides which statistically differ between pre-menarcheal and post-menarcheal Caucasian girls consuming a low soy diet, then use nanoLC-MALDI and tandem MS to identify the proteins from which the peptides were derived, and finally develop nanoLC-MRM-tandem MS methods to quantitatively assay the puberty-associated peptides;(2) using the established procedure, to evaluate whether age, time to or since menarche, and race/ethnicity impact these markers;to validate these markers using longitudinally collected specimens on individual girls going through puberty, taking into account any differences by age, time to or since menarche, or race/ethnicity identified in this aim;and (3) to determine if a high soy diet impacts pubertal development, as measured using these peptide markers, taking into account any confounding by the factors, and to determine if the levels of these peptide markers differ by level of soy consumption in pre-menarcheal and post-menarcheal girls. Project Narrative: The goal of this study is to define the stages of onset of puberty by applying modern proteomics methods to identify peptides in urines collected before and after the onset of menarche from a cohort of Caucasian and Asian girls. Longitudinal studies will allow the investigators to distinguish age-related change changes that are distinct from puberty itself. The effects of dietary intake of soy isoflavones and of race/ethnicity on the peptide markers will also be examined.